Bacteria Structure And Function

Question 1

3 domains of life

Answer 1

Prokaryotes
Archaea
Eukaryotes

Question 2

Structural differences between prokaryotes and eukaryotes

Answer 2

Size: prokaryotes smaller (50x smaller)
Nucleus: prokaryotes don’t have a nucleus
Organelles: prokaryotes don’t have organelles
Mitochondria: Prokaryotes lack mitochondria and instead produce their ATP on their cell surface membrane.
Peptidoglycan cell wall: present in prokaryotes
Ribosomes: ribosomes smaller in prokaryotes (70S) [80S in eukaryotes]
DNA: singular circular genome with no histones (prokaryotes) [eukaryotes have multiple linear chromosomes (histones)]

Question 3

How do you measure bacterial growth in liquid media (broth)

Answer 3

Measure cell density of broth
- called measuring the optical density if the broth

Question 4

What is lag phase

Answer 4

Bacteria are taking a while to divide

Question 5

What is exponential growth phase

Answer 5

Bacteria are replicating at their maximal rate

Question 6

What is stationary phase

Answer 6

Bacteria run out of nutrients (use up all the nutrients in the medium) so bacteria are replicating at the same rate that bacteria are dying

Question 7

Why can’t you pick up bacterial death when measuring optical (cell density)

Answer 7

Because a dead cell and alive cell will give the same broth density

Question 8

How to get a viable cell count

Answer 8

Take some of the broth (liquid medium with bacteria) and add it to an agar plate
You will see bacterial colonies

Question 9

How to measure viability (how many live bacteria in liquid broth)

Answer 9

Use the term CFC - colony forming unit

The number of cells originally in the broth is the same as the number of colonies that you see with naked eye

Question 10

What do bacteria need to grow

Answer 10

Correct temp
Correct pH
Carbon source (can be breakdown of lipids, proteins)
Oxygen or No oxygen
Potassium, magnesium, calcium and cofactors

Question 11

Basic difference between gram + and gram - bacteria

Answer 11

All bacteria have cytoplasmic membrane phospholipid bilayer

Gram negative cells have a thin peptidoglycan layer

And an outer membrane

Gran positive bacteria have a thick peptidoglycan

But no outer membrane

Question 12

Function of cell envelope

Answer 12

Deal with stress

Question 13

How do you tell the difference between gram + and gram - cell walls (in a lab)

Answer 13

Gram stain
Gram positive bacteria will stain purple
Gram negative bacteria will stain pink

Question 14

How to perform a bacterial gram stain

Answer 14

Prepare a bacterial smear on a clean microscope slide.
This is done by adding a small amount of sterile water to the bacterial colony and spreading it evenly over the slide.

Allow it to air dry.

Heat fix the bacterial smear by passing the slide through a flame 2-3 times. This will kill the bacteria and help them adhere to the slide.

Flood the bacterial smear with crystal violet stain and let it sit for 1 minute.
Rinse the slide gently with water to remove excess stain.

Add Gram’s iodine solution to the slide and let it sit for 1 minute. This will act as a mordant to fix the crystal violet stain to the cell walls.

Rinse the slide gently with water to remove excess iodine.

Decolorize the slide by adding 95% ethanol dropwise until the runoff is clear.
This step is critical, as over-decolorization can lead to false-negative results, while under-decolorization can lead to false-positive results.

Rinse the slide gently with water to remove excess ethanol.

Counterstain the slide by adding safranin stain and let it sit for 1 minute. This will stain any remaining bacteria that were not stained by the crystal violet.

Rinse the slide gently with water to remove excess stain.
Blot the slide gently with a paper towel to remove excess water and let it air dry.

Place a cover slip over the bacterial smear and examine the slide under a microscope.

Gram-positive bacteria will appear purple, while Gram-negative bacteria will appear pink.

Question 15

Why does gram positive bacteria appear purple

Answer 15

Gram-positive bacteria appear purple after a Gram stain because they have a thick layer of peptidoglycan in their cell walls that traps the crystal violet-iodine complex during the staining process. The crystal violet-iodine complex is a purple dye that binds to the peptidoglycan layer of the cell wall, making the bacteria appear purple under a microscope.

After decolorization with ethanol, the thick peptidoglycan layer of the Gram-positive bacteria retains the crystal violet-iodine complex, while the thinner peptidoglycan layer of Gram-negative bacteria does not. This leads to the differential staining pattern where Gram-negative bacteria appear pink after counterstaining with safranin, while Gram-positive bacteria retain their purple color.

Question 16

Why do Gram-negative bacteria not retain the violet dye in a Gram stain

Answer 16

Gram-negative bacteria do not retain the violet dye in a Gram stain because they have a thinner layer of peptidoglycan in their cell walls compared to Gram-positive bacteria.

During the Gram stain procedure, the violet dye (crystal violet) is first added to the bacterial sample, which stains all bacteria in the sample purple. Then, a decolorizing agent (usually alcohol or acetone) is added to the sample, which washes away the violet dye from the thinner peptidoglycan layer of Gram-negative bacteria.

The thinner peptidoglycan layer of Gram-negative bacteria makes them more susceptible to the action of the decolorizing agent, which removes the violet dye from their cell walls. Following decolorization, the Gram-negative bacteria are stained pink or red by the counterstain (usually safranin), which is used to visualize them.

Question 17

What is peptidoglycan

Answer 17

An amino sugar

Question 18

Chemistry of peptidoglycan

Answer 18

It is a complex polymer made up of long chains of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) molecules cross-linked by short peptide chains. The peptide chains are composed of amino acids such as L-alanine, D-alanine, D-glutamic acid, and L-lysine.

Question 19

Differences between gram positive and gram negative bacteria (detailed)

Answer 19

Both have cell membrane - phospholipid bilayer

Gram positive
- thick peptidoglycan layer (begins straight from cell membrane)
- no lipopolysaccharides
- they do have other polysaccharides
- they have WTA - wall teichoic acids
- and they have LTAs - lipid teichoic acids
- WTA & LTA are different as WTA is loosely linked to peptidoglycan wall/layer, LTA is linked to outer leaflet of cytoplasmic membrane

Gram negative
- peptidoglycan later much thinner
- on top of peptidoglycan layer there is the outer membrane
-the space between the outer membrane and cell membrane (where the peptidoglycan layer is found) is called the periplasm
- outer membrane - bottom part is made of phospholipid - top part is made of lipopolysaccharide
- outer membrane is called an asymmetric bilayer because the outer leaflet isn’t the same as the inner leaflet
- different bacteria/bacterial strains will have different lipopolysaccharides on the outer layer

Question 20

Teichoic acid structure

Answer 20

Ribitol and glycerol linked by phosphate bonds

Question 21

Teichoic acid function

Answer 21

Maintaining cell shape and structural integrity: TAs are covalently linked to the peptidoglycan layer of the cell wall and contribute to its strength and stability. They also help to maintain the overall shape of the bacterial cell.

Regulating membrane permeability: TAs can modulate the permeability of the cell membrane, regulating the flow of molecules in and out of the cell.

Adhesion and colonization: TAs can play a role in bacterial adhesion to host cells and tissues, promoting colonization and infection.

Immune evasion: TAs can contribute to bacterial resistance to host immune responses by masking cell surface antigens or inducing the production of antibodies that are ineffective against the bacterium.

Question 22

Why do some bacteria have capsules

Answer 22

Protection from the host immune system: Capsules can prevent recognition and phagocytosis by host immune cells, making it difficult for the host to clear the infection. This is particularly important for bacterial pathogens that need to evade the host immune response to establish infection.

Attachment to surfaces: Capsules can help bacteria adhere to surfaces, which is important for forming biofilms, colonizing host tissues, and establishing infections.

Resistance to desiccation and environmental stress: Capsules can help bacteria resist desiccation and other environmental stresses, which can be especially important for bacteria that live in dry or nutrient-poor environments.

Resistance to antibiotics: Capsules can help bacteria resist the effects of antibiotics, making them more difficult to treat.

Question 23

What are capsules

Answer 23

Capsules are outermost layers of polysaccharides that surround some bacterial cells.

Question 24

Difference between fimbriae and pili

Answer 24

While the fimbriae are bristle-like short fibers occurring on the bacterial surface, Pili are long hair-like tubular microfibers found on the surface of bacteria.

Question 25

What is the function of fimbriae and pili

Answer 25

Attachment to surfaces: Fimbriae and pili are important for bacterial attachment to surfaces, including host tissues, which is critical for the establishment of infections.
Biofilm formation: Fimbriae and pili are important for the formation of bacterial biofilms, which are communities of microorganisms that are embedded in a matrix of extracellular polymeric substances.
Motility: Some types of fimbriae and pili are involved in bacterial motility, including twitching and gliding motility, which allows bacteria to move across surfaces.
Conjugation: Some types of pili are involved in bacterial conjugation, which is a mechanism of horizontal gene transfer that allows bacteria to exchange genetic material.

Pili also involved in genetic exchange

Question 26

Flagella function

Answer 26

Motility